Abstract

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Natural dibenzo-α-pyrones are an important group of metabolites derived from fungi, mycobionts, plants and animal feces. They exhibit a variety of biological activities such as toxicity on human and animals, phytotoxicity as well as cytotoxic, antioxidant, antiallergic, antimicrobial, antinematodal, and acetylcholinesterase inhibitory properties. Dibenzo-α-pyrones are biosynthesized via the polyketide pathway in microorganisms or metabolized from plant-derived ellagitannins and ellagic acid by intestinal bacteria. At least 53 dibenzo-α-pyrones have been reported in the past few decades. This mini-review aims to briefly summarize the occurrence, biosynthesis, biotransformation, as well as their biological activities and functions. Some considerations related to synthesis, production and applications of dibenzo-α-pyrones are also discussed.

1. Introduction

Dibenzo-α-pyrones (also named dibenzo-α-pyranones, 6H-benzo[c]chromen-6-ones, and 6H-dibenzo[b,d]pyran-6-ones) are an important group of heptaketide coumarin derivatives that have a fused tricyclic nucleus (Figure 1). They are usually isolated from fungi [1], mycobionts [2,3], plants [4,5], and animal feces containing the transformed products of plant-derived ellagitannins and ellagic acid by intestinal bacteria [6,7]. Many of them possess a wide spectrum of biological activities, spanning from toxicity on human and animals [8], cytotoxic activity [9], phytoxicity [10], antioxidant [6], antiallergic [11], antimicrobial [12], to acetylcholinesterase inhibitory activities [13]. In addition, dibenzo-α-pyrones are key intermediates in the synthesis of cannabinoids [14], and other pharmaceutically interesting compounds such as progesterone, androgen, glucocorticoid receptor agonists [15,16], as well as endothelial proliferation inhibitors [17], and antidyslipidemic agents [18]. This review mainly presents the occurrence, biosynthesis, biotransformation, and biological activities of the dibenzo-α-pyrones from bioorganisms. We also discuss and prospect their synthesis, production and applications.

2. Occurrence

2.1. Dibenzo-α-pyrones from Fungi

Dibenzo-α-pyrones are mainly distributed in the Alternaria species and mycobionts. Other dibenzo-α-pyrone-producing fungi include Botrytis allii, Cephalosporium acremonium, Hyalodendriella sp. Ponipodef12, Microsphaeropsis olivacea, Penicillium verruculosum, and Phoma sp. TC 1674 (Table 1). From the biosynthetic pathway, fungal dibenzo-α-pyrones have a polyketide origin via acetyl-CoA and malonyl-CoA. They are usually toxic to plants and animals. Typical examples include alternariol (10), alternariol 9-methyl ether (11), botrallin (16), and 2-chloro-4,6-dihydro-1,7-dihydroxy-3,9-dimethoxy-1-methyl-1H-dibenzo[b,d]pyran-4,6-dione (TMC-264, 28). The structures of the dibenzo-α-pyrones from fungi are shown in Figure 2.

2.2. Dibenzo-α-pyrones from Plants

The dibenzo-α-pyrones from plants are listed in Table 2. One dibenzo-α-pyrone, namely djalonensone (11), was isolated from the roots of Anthocleista djalonensis (Loganiaceae). The authors postulated djalonensone to be a significant taxonomic marker of the plant species [48]. However, djalonensone is identical to alternariol 9-methyl ether (AME) which has been isolated from a series of fungi including pathogenic and endophytic fungi [1]. Thus, the significance of djalonensone (11) as an important taxonomic marker of the plant species should be reconsidered. The possibility that djalonensone (11) was produced by an endophytic fungus residing in the healthy roots of A. djalonensis, needs further investigation [49].

Three dibenzo-α-pyrones, namely (2'S,3'R)-3,10-dihydroxy-9-O-(6'-hydroxy-2'-hydroxymethyl- dihydrofuran-3-yl)-dibenzo[b,d]pyran-6-one (31), (2'S,3'R)-3,10-dihydroxy-9-O-(5',6'-dihydroxy-2'-hydroxymethyldihydrofuran-3-yl)-dibenzo[b,d]pyran-6-one (32), and fasciculiferol (33) were isolated from the heartwood of Umtiza listerana [50]. Fasciculiferol (33) was previously isolated from the heartwood of Acacia fasciculifera [51]. Two dibenzo-α-pyrones, autumnariniol (29) and autumnariol (30), were isolated from the bulbs of Eucomis autumnalis (Liliaceae) [52]. Four urolithins, namely urolithin A (40), isourolithin A (41), urolithin B (42) and urolithin C (43) were isolated from Trapa natans (Trapaceae) [4] and Caesalpinia sappan (Caesalpiniaceae) [53]. These urolithins (Figure 3) were also isolated from animal feces [6,7]. Other dibenzo-α-pyrones isolated from plants included sabilactone (38) from Sabina vulgaris [54] and sarolactone (39) from Hypericum japonicum [55]. The structures of the dibenzo-α-pyrones from plants are shown in Figure 4.

2.3. Dibenzo-α-pyrones Produced by Transformation of Intestinal Bacteria

A group of dibenzo-α-pyrones 40–52, namely urolithins with different phenolic hydroxylation patterns, have been isolated from animal feces. Ellagitannins and ellagic acid (EA) are plant secondary metabolites that have relevant antioxidant activities in vitro, potential cardiovascular protection, anticarcinogenic and anti-inflammatory effects [59,60,61]. These dibenzo-α-pyrones are important constituents in different foods including pomegranates, berries (i.e., strawberry, raspberry, blackberry, and camu-camu), nuts (i.e., walnuts, acorns, and chestnuts), muscadine grapes, oak-aged wines, medicinal plants and tisanes (i.e., geranium and oak leaves). They are not absorbed in the gut and are metabolized in vivo by the intestinal bacteria to produce a series of metabolites known as urolithins [62,63]. Some urolithins such as urolithins A (40), B (42) and C (43) as well as isourolithin A (41) were previously isolated from plants (Table 2) [4,53]. The structures of the isolated urolithins are shown in Figure 3.

2.4. Dibenzo-α-pyrones from Bacteria

Up to now, only one dibenzo-α-pyrone called murayalactone (53) has been isolated from Streptomyces murayamaensis [64]. The structure of murayalactone is shown in Figure 5.

3. Biosynthesis and Biotransformation

We know very little about the biosynthesis of dibenzo-α-pyrones in living organisms, including their genetics, biochemistry and biosynthetic pathways [2,25,65,66]. In plants, gallic acid (54), which is biosynthesized via the shikimate pathway [67], was considered as the precursor of ellagitannins [68]. The ellagitannins would be transformed into ellagic acid (55), and then into a series of urolithins (Scheme 1) [6,7,63]. However, in microorganisms, dibenzo-α-pyrones are biosynthesized via the polyketide pathway [3,25]. Polyketide synthase (PKS) is one of the postulated core enzymes in the biosynthesis of 6H-dibenzo[b,d]pyran-6-ones (i.e., alternariol, AME) in Alternaria alternata [66]. In a draft genome sequence of A. alternata, 10 putative PKS-encoding genes were identified. The timing of the expression of two PKS genes, pksJ and pksH, was found to correlate with the production of AOH and AME [66].

Alternariol (10) was first thought to be biosynthesized via norlichexanthone [65], which was ruled out later, and it was proven that alternariol (10) could be biosynthesized by simple cyclization and aromatization of a polyketide precursor [69]. After administration of [1-13C]acetate and [1,2-13C]acetate to cultured lichen mycobionts of Graphis spp., acetate units were incorporated into the 6H-dibenzo[b,d]pyran-6-one derivatives including alternariol (10), AME (11) and graphislactones A-F (18–23) [3]. Alternariol (10) could suffer oxidative demethylation at C-1, hydroxylation at C-4, and O-methylation at C-9 to lead formation of graphislactones E (22) and F (23). On the other hand, alternariol (10) could be transformed to graphislactones A-D (18–21) without demethylation via AME (11) [3]. The biosynthetic pathways of graphislactones A-F (18–23) in the cultured lichen mycobionts of Graphis spp. are shown in Scheme 2.

Epigenetic modifiers, including DNA methyltransferase (DNMT) inhibitors (i.e., 5-azacytidine, abbreviated as 5-AC) and histone deacetylase (HDAC) inhibitors (i.e., suberoylanilide hydroxamic acid, abbreviated as SBHA) are useful to induce the expression of otherwise silent biosynthetic genes under standard laboratory conditions [70]. Supplementation of a DNMT inhibitor 5-AC or a HDAC inhibitor SBHA to the medium induced the production of alternariol (10), alternariol 9-methyl ether (11), 4-hydroxyalternariol 9-methyl ether (14) and altenusin (56) [25]. The proposed biosynthesis of the aromatic polyketides 10, 11, 14 and 56 involves the condensation of seven molecules of malonyl-CoA, followed by aldol-type cyclizations between C-2 and C-7, and C-8 and C-13, and the subsequent lactonization leads to alternariol (10) (Scheme 3). On the other hand, subsequent methylation of the C-9 hydroxyl group of alternariol (10) by a methyltransferase results in 4-hydroxyalternariol 9-methyl ether (14). Furthermore, the reduction of the C-9 carbonyl group of the heptaketide intermediate by a reductase, and subsequent aldol-type cyclization would produce a biphenyl. Methylation of the C-5 hydroxyl group, and the hydroxylation of C-5' would then lead to altenusin (56). The hypothetical biosynthetic pathways [25] of alternariol (10) and its derivatives 11, 14 in an endophytic fungus from Datura stramonium are shown in Scheme 3.

Urolithins include a family of metabolites of dibenzo-α-pyrone structures with different phenolic hydroxylation patterns. They are produced in different animals after the intake of ellagitannins and ellagic acid (EA) [71,72]. Ellagitannins are hydrolyzed to ellagic acid (55) in the acidic environment of the stomach by the action of the intestinal bacteria. The proposed transformation from ellagic acid to urolithins by the intestinal bacteria [6,7,63] is shown in Scheme 1.

4. Biological Activities and Functions

Dibenzo-α-pyrones and their derivatives with diverse chemical properties have been clarified (Figure 1, Figure 2, Figure 3, Figure 4 and Figure 5, Table 1 and Table 2). Some of them act as mycotoxins to humans and animals or as phytotoxins to plants. They have been examined to have a variety of biological activities and functions, which mainly include the cytotoxic, antioxidant, antiallergic, antimicrobial, antinematodal, and acetyl-cholinesterase inhibitory activities.

4.1. Toxicity on Human and Animals

The association of mycotoxins from Alternaria fungi with human and animal health is not a recent phenomenon. Alternaria toxins have been linked to a variety of adverse effects (i.e., genotoxic, mutagenic, and carcinogenic) on human and animal health [8]. Altenuene (1), alternariol (10), and alternariol 9-methyl ether (11) were studied for their toxicity to chickens. Addition of these compounds in chicken feed from sublethal to lethal levels progressively reduced feed efficiency, suppressed weight gain and increased internal haemorrhaging [27,73].

There were a few reports about the toxicity of Alternaria metabolites on brine shrimp (Artemia salina L.) [74,75]. The LC50 values of altenuene (1) and alternariol (10) were 375 and 100 µg/mL, respectively, to brine shrimp larvae by using the disk method of inoculation and an exposure period of 18 h [75]. Altenuene (1) and alternariol (10) along with alternariol 9-methyl ether (11) were also verified to be toxic to brine shrimp [74].

4.2. Cytotoxic Activity

Among Alternaria dibenzo-α-pyrones, alternariol (10) was the most active metabolite to have cytotoxic activity on L5178Y mouse lymphoma cells [9], as well as to have inhibitory activity on protein kinase and xanthine oxidase [28]. Further investigation showed that alternariol (10) was a topoisomerase I and II poison which might contribute to the impairment of DNA integrity in human colon carcinoma cells [73,76]. It induced cell death by activation of the mitochondrial pathway of apoptosis in human colon carcinoma cells [76]. Alternariol (10) and its 9-methyl ether (11) induced cytochrome P450 1A1 and apoptosis in murine heptatoma cells dependent on the aryl hydrocarbon receptor [77]. Other alternariol derivatives such as alternariol 9-methyl ether (11), alternariol 9-O-sulfate (13), and altenusin (56) were also screened to be cytotoxic [9].

Dehydroaltenusin (17), isolated from A. tenuis, was found to be a specific inhibitor of eukaryotic DNA polymerase α to show its strong cytotoxic activity on tumor cells [45,78]. This compound also exhibited strong inhibitory activity on mammalian DNA polymerase α in vitro [79]. It was further proved to abrogate cell proliferation of the cultured mammalian cells to show its potential as an effective chemotherapeutic agent against tumors [44].

4.3. Phytotoxicity

The metabolites from fungal pathogens are usually toxic to plants and are called phytotoxins which are divided into host-specific [83,84] and host non-specific toxins [85,86]. Some Alternaria-derived dibenzo-α-pyrones were approved as the host-specific phytotoxins including altenuene (1), alternariol (10), alternariol 9-methyl ether (11), alternuisol (15), and dehydroaltenusin (17) [9,10,37,39,41,45].

4.4. Antioxidant Activity

Urolithin A (40), isourolithin A (41), and urolithin B (42) from the fruits of Trapa natans showed antioxidant activity. Among them, isourolithin A (41) showed the strongest, and urolithin B (42) showed weak antioxidative effect [4]. As ellagic acid and ellagitannins are extremely poorly absorbed in gut, urolithins appear to be responsible for biological activities related to the intake of ellagitannins. Most of urolithins (i.e., urolithins A, C, and D) exhibited antioxidant activity in a cell-based assay [6]. However, there have been contradictory reports on their antioxidant capacity [62,87]. Recently, urolithins were revealed to display both antioxidant and pro-oxidant activities depending on assay system and conditions by using oxygen radical absorbance capacity (ORAC) assay, three cell-based assays, copper-initiated pro-oxidant activity (CIPA) assay, and cyclic voltammetry. Urolithins were screened to be the strong antioxidants in the ORAC assay, but mostly pro-oxidants in cell-based assays [88]. The antioxidant activity of urolithins is very likely mediated exclusively by the hydrogen atom transfer (HAT) mechanism. The hydrogen atom is donated by the phenolic hydroxyl group [88].

4.5. Antiallergic Activity

Urolithin A (40), isourolithin A (41), and urolithin B (42), from the feces of Trogopterus xanthipes showed hyaluronidase inhibitory activities with IC50 values of 1.33, 1.07 and 2.33 mM, respectively that indicated their antiallergic activity [11]. TMC-264 (28) from the fungus Phoma sp. TC 1674 [47] selectively inhibited tyrosine phosphorylation of STAT6, and also inhibited the complex formation of phosphorylated STAT6 and its recognition sequence. Therefore, TMC-264 (28) would inhibit IL-4 signaling and would be useful in the treatment of allergic disease [47,89,90].

Both urolithins A (40) and B (42) from human feces exhibited estrogenic and antiestrogenic activities, which suggested that consumption of ellagitannin-containing foodstuffs such as pomegranate, walnuts, berries, and oak-aged wines may exert some proestrogenic/antiestrogenic effects [91].

5. Conclusions and Future Perspectives

We have just clarified one part of the dibenzo-α-pyrones from fungi, plants and bacteria. The remaining dibenzo-α-pyrones in bioorganisms need to be further identified. In recent years, more and more dibenzo-α-pyrones have been isolated from plant endophytic fungi. These endophytic fungi could be the rich sources of biologically active compounds that are indispensable for medicinal and agricultural applications [92,93]. In most cases, biological activities, structure-activity relationships, and modes of action of dibenzo-α-pyrones were only primarily investigated.

With comprehensive understanding of the biosynthetic pathways of some dibenzo-α-pyrones in the next few years, we may be able to effectively not only increase the yields of bioactive dibenzo-α-pyrones, but also block the biosynthesis of some toxic dibenzo-α-pyrones (i.e., phytotoxins and mycotoxins) [1].

Acknowledgements

This work was co-financed by the grants from the National Basic Research Program of China (2010CB126105), the Hi-Tech R&D Program of China (2011AA10A202), and the National Natural Science Foundation of China (31271996).